Method of substance separation by supercritical fluid chromatography and vapor liquid separator for use therein

ABSTRACT

A method comprising liquefying a gas to thereby obtain a liquefied gas; injecting a sample into a mobile phase containing a solvent and a supercritical fluid formed from the liquefied gas; passing this mobile phase through a column so that the mobile phase containing desired substance is divided into the solvent and the gas; and separating the desired substance from the solvent, wherein when the pressure of the gas divided from the mobile phase is higher than the pressure of the gas fed for formation of the liquefied gas from gas supply device, the gas divided from the mobile phase is liquefied. Further, there is provided a vapor liquid separator for use in the method.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a divisional of U.S. application Ser. No. 11/478,699filed Jul. 3, 2006 now U.S. Pat. No. 7,678,276, which is a continuationof PCT International Application No. PCT/JP2004/019105 filed on Dec. 21,2004, which designated the United States and upon which priority isclaimed under 35 U.S.C. §120. This application also claims priorityunder 35 U.S.C. §119(a) on Patent Application No(s). 2004-000317 filedin Japan on Jan. 5, 2004. The entire contents of each of the aboveapplications is hereby incorporated by reference.

TECHNICAL FIELD

The present invention relates to a method of substance separation bysupercritical fluid chromatography of recovering a gas componentconstituting a supercritical fluid from a mobile phase containing thesupercritical fluid and a solvent and recycling the gas component, and avapor liquid separator for use in the method.

BACKGROUND ART

Each of various kinds of chromatography has been used as a method ofseparating a desired substance in a sample. A known example of such thechromatography is supercritical fluid chromatography using asupercritical fluid as a mobile phase. Since the supercritical fluidchromatography uses a fluid, which is referred to as a supercriticalfluid and has a greater variety of properties than that of a generalsolvent, as a mobile phase, investigation has been conducted into theutilization of supercritical fluid chromatography for the separation,analysis, purification, and the like of various substances that havebeen considered to be difficult to separate.

A technique has been known as the method of substance separation bysupercritical fluid chromatography, which involves: delivering asupercritical fluid supplied from a bomb by using a pump; injecting asample into the supercritical fluid; passing the supercritical fluidinto which the sample has been injected through a column; reducing thepressure of the supercritical fluid that has been passed through thecolumn to precipitate a solute; dividing the precipitated solute and thesupercritical fluid; and delivering the divided supercritical fluid tothe pump to recycle the supercritical fluid (see, for example, JapanesePatent Laid-Open Publication 05-307026).

By the way, a mobile phase obtained by mixing a supercritical fluid anda solvent is used in supercritical fluid chromatography depending on thekind of a solute and the kind of a filler in some cases. The abovetechnique does not show the use of such mobile phase, so the recyclingof the supercritical fluid upon use of such mobile phase is susceptibleto investigation.

DISCLOSURE OF THE INVENTION

The present invention is to provide a method of separating a substanceby supercritical fluid chromatography using a mobile phase containing asupercritical fluid and a solvent, the method allowing the recycling ofa gas from which the supercritical fluid is formed, and a vapor liquidseparator for use in the method.

The present invention provides a method including the steps of: forminga supercritical fluid from a gas; subjecting a mobile phase that hasbeen passed through a column to vapor liquid separation; setting thepressure of the divided gas to be higher than a pressure of a gas to benewly supplied; and placing higher priority on the use of the dividedgas for the formation of the supercritical fluid than on the use of thegas to be newly supplied for the formation when the pressure of thedivided gas is higher than the pressure of the gas to be newly supplied,and a vapor liquid separator to be suitably used for the method.

In other words, the present invention relates to a method of separatinga substance by supercritical fluid chromatography comprising the stepsof: liquefying a gas to form a liquefied gas; injecting a samplecomprising a desired substance into a mobile phase comprising asupercritical fluid formed from the resultant liquefied gas and asolvent; passing the mobile phase into which the sample has beeninjected through a column for separating the desired substance from thesample; and dividing the mobile phase comprising the desired substanceinto the solvent and the gas; so that the desired substance is separatedfrom the solvent, wherein, in the step of forming a liquefied gas, whena pressure of the gas divided from the mobile phase is higher than apressure of a gas supplied for formation of the liquefied gas from a gassupply device, the gas divided from the mobile phase is liquefied(hereinafter, the method of the present invention is referred to“separation method” simply).

In addition, the present invention relates to a vapor liquid separatorfor use in a method of separating a substance by supercritical fluidchromatography of injecting a sample containing a desired substance intoa mobile phase comprising a supercritical fluid and a solvent, passingthe sample through a column for separating the desired substance fromthe sample to separate the desired substance in the sample, dividing themobile phase that has been passed through the column into the solventand a gas, and separating the desired substance from the solvent. Thevapor liquid separator comprises: an external cylinder having ends bothof which being opened; a flange portion for clogging an opening at oneend of the external cylinder; an introduction portion for introducingthe mobile phase into the external cylinder in a circumferentialdirection along an inner peripheral wall surface of the externalcylinder; and an internal cylinder having ends both of which beingopened and which penetrates through the flange portion to extend closerto an other end side of the external cylinder than the introductionportion.

BRIEF DESCRIPTION OF THE DRAWINGS

[FIG. 1] A schematic view showing the structure of an example of asupercritical fluid chromatographic fractionation apparatus for use in aseparation method of the present invention.

[FIG. 2] A sectional view showing the vertical section of a main portionof an example of a vapor liquid separator of the present invention.

[FIG. 3] A sectional view showing the cross section of a main portion ofthe vapor liquid separator shown in FIG. 2 taken along the line A-A.

BEST MODE FOR CARRYING OUT THE INVENTION

<Method of Separating a Substance by Supercritical Chromatography>

A separation method of the present invention comprises the steps of:liquefying a gas to form a liquefied gas; injecting a sample containinga desired substance into a mobile phase containing a supercritical fluidformed from the resultant liquefied gas and a solvent; passing themobile phase into which the sample has been injected through a columnfor separating the desired substance from the sample; and dividing themobile phase containing the desired substance into the solvent and thegas.

In the step of forming a liquefied gas, the gas from which thesupercritical fluid is formed is liquefied once. Such formation of theliquefied gas is performed by regulating the pressure and temperature ofthe gas. A pressure regulating device such as a compressor or a backpressure regulating valve can be used for regulating the pressure of thegas. Temperature regulating device such as a heat exchanger can be usedfor regulating the temperature of the gas.

Examples of the above-mentioned gas which can be used herein includecarbon dioxide, ammonia, sulfur dioxide, hydrogen halide, nitrous oxide,hydrogen sulfide, methane, ethane, propane, butane, ethylene, propylene,halogenated hydrocarbon, and water. Of those, carbon dioxide ispreferable in view of combustibility, explosivility, harmful nature tohumans, ease of handling, cost efficiency, and the like.

The mobile phase in the step of injecting a sample is a mixed solventcomprising a supercritical fluid and a solvent. The supercritical fluidin the present invention is the gas having at least one or both of apressure exceeding a critical pressure and a temperature exceeding acritical temperature. The supercritical fluid can be formed by furtherapplying one or both of a pressure and heat to the formed liquefied gas.The mobile phase may be formed by mixing the formed supercritical fluidand the solvent, or may be formed by: mixing the liquefied gas and thesolvent; applying one or both of a pressure and heat to the mixedsolvent; and turning the liquefied gas in the mixed solvent into asupercritical fluid.

As performed in, for example, high performance liquid chromatography,the mixing of the liquefied gas or the supercritical fluid with thesolvent can be performed by delivering each of fluids by using ametering pump at a predetermined flow rate and by mixing the fluids. Themixing ratio of the solvent, which varies depending on conditions suchas a kind of a substance in the sample, is preferably about 5 to 30% bymass in the mobile phase for improving the efficiency of the separationof the desired substance. A known solvent to be selected in accordancewith, for example, the kind of a substance to be separated and the kindof a filler is used as the solvent. Examples of the solvent to be usedinclude lower alcohols such as ethanol and 2-propanol.

Known means such as an injector ordinarily used in high performanceliquid chromatography can be used for injecting the sample into themobile phase. A sample obtained by means of an ordinary method involvingdissolving of a separation object into an appropriate solvent can beused as the solvent. Any one of general organic solvents or a mixedsolvent of the solvents can be used as the solvent for dissolving theseparation object. An injection method causing no fluctuation inpressure upon injection as described in Japanese Patent Laid-OpenPublication 05-307026 mentioned above is preferably adopted in injectingthe sample for improving the efficiency of the separation of the desiredsubstance.

An appropriate column in accordance with the desired substance is usedin the step of passing the mobile phase into which the sample has beeninjected through a column. The present invention can be suitablyutilized for the separation of a substance that has been difficult toseparate by any other kind of chromatography because the presentinvention uses a mobile phase containing a supercritical fluid. Forexample, when the sample contains a mixture of optical isomers and oneattempts to separate one of the isomers, a column filled with a fillerobtained by causing a carrier such as silica to carry a polysaccharidederivative having an optical isomer separating ability on its surface isused as the column.

An ordinary vapor liquid separator capable of dividing a solvent and agas can be used in the step of dividing the mobile phase containing thedesired substance into the solvent and the gas; however, a vapor liquidseparator of the present invention to be described later is preferablyused for performing high-accuracy, stable division. The desiredsubstance can be obtained from the solvent by: releasing the dividedsolvent from a high-pressure condition; and adopting a known method suchas vacuum concentration as required.

In the present invention, the gas divided from the mobile phase isrecycled as a raw material for the liquefied gas in accordance with thepressure of the gas. That is, in the step of forming a liquefied gas,when the pressure of the gas divided from the mobile phase is higherthan a pressure of a gas supplied for the formation of the liquefied gasfrom a gas supply device, the gas divided from the mobile phase isliquefied.

Such recycling of the divided gas can be performed by: connecting thevapor liquid separator and a device for forming the liquefied gas byusing a pipe; and setting the pressure of the divided gas to be higherthan the pressure of the gas from the gas supply device by usingappropriate pressure regulating device. As described above, a knowndevice such as a compressor or a back pressure regulating valve can beused as the pressure regulating device.

The pressure of the divided gas is set in accordance with not only thepressure of the gas from the gas supply device but also the solubilityof the gas in the solvent from the viewpoint of the suppression of theredissolution of the divided gas into the solvent. For example, whencarbon dioxide is used as the gas and an ordinary organic solvent isused as the solvent, the pressure of the divided gas is preferably setto about 1 to 10 MPa for suppressing the redissolution of the dividedgas into the solvent.

The pressure of the divided gas is not particularly limited so long asit is set to be higher than the pressure of the gas from the gas supplydevice; however, a difference between the pressure of the divided gasand the pressure of the gas from the gas supply device is preferably 0.1MPa or more.

The present invention may further include the step of dividing the gasfrom the solvent that has been divided in the dividing step. Suchredividing step enables an increased amount of the gas to be recoveredfrom the solvent, and can further increase the recovery rate of the gas.

The redividing step can be performed by reducing the pressure of asystem storing the solvent. The pressure of the system storing thesolvent can be continuously reduced by, for example, storing the solventin a hermetically sealable container and gradually opening thehermetically sealable container to a lower pressure atmosphere such as acontainer with normal pressure or a vacuum pump.

The redividing step can be performed also by storing the solvent in ahermetically sealable container and setting the pressure of thehermetically sealable container to be lower than the pressure of thesolvent before the storage. The pressure in the container may be setbefore the storage of the solvent, or may be set after the storage ofthe solvent.

Furthermore, the redividing step can be performed by connecting multiplehermetically sealable containers of the above kind in series andsequentially supplying the solvent to a container with a reducedinternal pressure. The use of the multiple hermetically sealablecontainers as mentioned above enables an operation for dividing the gasand the solvent from the mobile phase by using the vapor liquidseparator and an operation for further recovering the gas from thesolvent to be performed in tandem with each other. The number of thehermetically sealable containers, which is not particularly limited, ispreferably about 1 to 3 from the viewpoints of the simplicity of afacility or operation, improvement in recovery rate of the gas from thesolvent, and the like.

The gas divided in the redividing step can be recycled as the liquefiedgas by, for example, recovering the gas from the hermetically sealablecontainer and compressing the recovered gas by using a compressor toliquefy the gas.

The redividing step is particularly effective in the case where a largeamount of the solvent is divided in the dividing step. An example ofsuch case is the case where a column with a large diameter is used forseparating the desired substance such as industrial production. Thediameter of the column in such case is preferably 10 cm or more, or morepreferably 20 cm or more.

The present invention may include any other step except those describedabove. Examples of the other step include: the step of detecting thesubstance in the mobile phase that has been passed through the column;the step of supplying the mobile phase to one of multiple vapor liquidseparation devices in accordance with the result of the detection of thesubstance in the mobile phase; and the step of purifying the dividedgas.

<Vapor Liquid Separator>

A vapor liquid separator of the present invention comprises: an externalcylinder having ends both of which being opened; a flange portion forclogging an opening at one end of the external cylinder; an introductionportion for introducing the mobile phase into the external cylinder in acircumferential direction along the inner peripheral wall surface of theexternal cylinder; and an internal cylinder having ends both of whichbeing opened and which penetrates through the flange portion to extendcloser to the other end side of the external cylinder than theintroduction portion.

According to the vapor liquid separator, the internal cylinder extendscloser to the other end side of the external cylinder than theintroduction portion, so a gas to be discharged from the internalcylinder hardly contains a droplet of the mobile phase introduced fromthe introduction portion, and the concentration of the solvent or of thesubstance in the gas can be reduced. In addition, the external cylinderis of a cylindrical shape, so a pressure is uniformly applied to theinner peripheral wall surface and the durability of the vapor liquidseparator is improved. As described above, the vapor liquid separatorcan recover a gas with a high purity under a high-pressure condition, soit is suitably used for a technique for recycling a recovered gas as amobile phase in the separation of optical isomers by supercritical fluidchromatography requiring improved accuracy.

For further improving separation efficiency in vapor liquid separationin the external cylinder, the vapor liquid separator preferably furthercomprises: a jacket that covers the outer peripheral wall surface of theexternal cylinder for regulating the temperature of the externalcylinder; and a mobile phase supply pipe that passes the inside of thejacket to be connected to the introduction portion for supplying themobile phase to the introduction portion, because the temperature of themobile phase is regulated to the temperature of the external cylinderbefore the mobile phase is introduced into the external cylinder.

The external cylinder is not particularly limited so long as the mobilephase introduced from the introduction portion flows from one end sideof the external cylinder to the other end side along the innerperipheral wall surface in a spiral fashion. The external cylinder,which is generally of a cylindrical shape, may be a cylinder having theinner peripheral wall surface of a substantially circular sectionalshape such as an elliptical shape or a polygonal shape for the purposeof, for example, causing a bias in the flow velocity of the mobile phaseintroduced into the external cylinder or applying an appropriate impact.

The flange portion is not particularly limited so long as it clogs anopening at one end of the external cylinder. For example, reducingunions for installing the internal cylinder and clogging the externalcylinder can be suitably used as such flange portion.

The introduction portion can be formed of a nozzle or pipe forintroducing the mobile phase into the external cylinder along thetangent of the inner peripheral wall surface in the sectional shape ofthe external cylinder. The mobile phase is ordinarily introduced in thedirection parallel with the transverse direction of the externalcylinder. How to provide the introduction portion is not limited to thatmentioned above in the present invention. The introduction portion maybe provided to be in parallel with the transverse direction of theexternal cylinder, or may be provided obliquely to the transversedirection.

The sectional shape of the internal cylinder is not particularly limitedso long as the internal cylinder is a pipe having a diameter smallerthan that of the external cylinder. The internal cylinder is constitutedto extend sufficiently closer to the other end side than theintroduction portion in the external cylinder from the viewpoint ofprevention of a mixing of a droplet of the mobile phase into a gas inthe external cylinder. The length by which the internal cylinder extendsvaries depending on various conditions such as conditions forintroducing the mobile phase into the external cylinder, the diameter ofthe external cylinder, and the diameter of the internal cylinder. Theinternal cylinder has only to be extended up to the position at whichthe flow of the mobile phase introduced into the external cylinder isprevented from losing its posture so that the mobile phase propagatesalong the inner peripheral wall surface of the external cylinder, thatis, a droplet is not generated from the inner peripheral wall surface.

An ordinary jacket for circulating a heat medium to the outside of theexternal cylinder can be used as the jacket. Using a hot heat medium forthe heat medium to be supplied to the jacket is effective in improvingthe efficiency of the division of the gas. Using a refrigerant for theheat medium to be supplied to the jacket is effective in reducing thesolubility of the desired substance in the solvent.

The mobile phase supply pipe is preferably provided over a wide range inthe jacket from the viewpoint of regulation of the temperature of themobile phase before vapor liquid separation to the temperature in theexternal cylinder in advance. For example, the mobile phase supply pipeis preferably provided in a meander shape in the jacket, or ispreferably provided in a spiral fashion for the outer peripheral wallsurface of the external cylinder. Such mobile phase supply pipe ispreferable for suppressing a droplet due to a change in temperature ofthe mobile phase upon introduction of the mobile phase into the externalcylinder particularly when the heat medium is a hot heat medium.

The vapor liquid separator of the present invention reduces a mixing ofa droplet of the mobile phase into the gas subjected to vapor liquidseparation. Accordingly, the vapor liquid separator is applicable toanalyzing and separating device using such gas as a mobile phase.Applying the vapor liquid separator to a device for analyzing andseparating optical isomers that may be greatly affected by mixing withimpurities is effective particularly in the field of, for example,medicine.

Hereinafter, an embodiment of the present invention will be described onthe basis of the drawings. First, FIG. 1 shows a supercritical fluidchromatographic fractionation apparatus for use in the separation methodof the present invention.

As shown in FIG. 1, the supercritical fluid chromatographicfractionation apparatus comprises: a bomb 1 as gas supply device filledwith carbon dioxide having a high pressure; a heat exchanger 2 forcooling and liquefying carbon dioxide having a high pressure; a pump 3for delivering the liquefied gas of carbon dioxide formed in the heatexchanger 2; a pump 5 for supplying a solvent supplied from a solventtank 4 to the liquefied gas delivered by the pump 3; a heat exchanger 6for heating a mixed solvent of the liquefied gas and the solvent to turnthe liquefied gas into a supercritical fluid; an injector 7 forinjecting a sample into a mobile phase as a mixture of the formedsupercritical fluid and the solvent; a column 8 for separating a desiredsubstance in the injected sample; a detector 9 for detecting thesubstance in the mobile phase that has passed the column 8; a backpressure regulating valve 10 as a pressure regulating device for keepingthe pressure in a system ranging from the pump 3 to the detector 9 at apredetermined pressure; multiple vapor liquid separators 11 eachintended to subject the mobile phase which has passed the back pressureregulating valve 10 and of which the pressure has been regulated tovapor liquid separation; tanks 12 each intended to store a liquid thathas been subjected to vapor liquid separation; a purifying device 13 forfurther removing a liquid from a gas that has been subjected to vaporliquid separation; a tank 14 for storing the liquid removed by thepurifying device 13; and a back pressure regulating valve 15 for keepingthe pressure of the gas purified by the purifying device 13 at apredetermined pressure.

The bomb 1, the heat exchanger 2, the pump 3, the heat exchanger 6, theinjector 7, the column 8, the detector 9, and the back pressureregulating valve 10 are connected in series by using pipes. The vaporliquid separators 11 are connected by using pipes in parallel with theback pressure regulating valve 10 and the purifying device 13. Thepurifying device 13 and the back pressure regulating valve 15 areconnected by using a pipe. The secondary side (side opposite to thepurifying device 13) of the back pressure regulating valve 15 isconnected to a pipe for discharging a gas to the outside of the system.The pipe for connecting the purifying device 13 and the back pressureregulating valve 15 bifurcates in midstream, and the bifurcated pipe isconnected to the pipe on the upstream side of the heat exchanger 2. Inaddition, the solvent tank 4 and the pump 5 are connected by using apipe. The pump 5 is connected to a pipe for connecting the pump 3 andthe heat exchanger 6 by using a pipe. Each of the vapor liquidseparators 11 and each of the tanks 12 are connected by using a pipe.The purifying device 13 and the tank 14 are connected by using a pipe.

A pressure regulating valve 16 for releasing carbon dioxide from thebomb 1 at a predetermined pressure and a check valve 17 for preventingthe back flow of a gas from the side of the heat exchanger 2 to the sideof the pump 1 are provided between the bomb 1 and the heat exchanger 2.A buffer tank 18 for receiving the liquefied gas formed in the heatexchanger 2 is provided between the heat exchanger 2 and the pump 3. Thecolumn 8 is stored in a column oven 19 for regulating the temperature inthe column 8 to be a predetermined temperature.

A valve 20 corresponding to each of the vapor liquid separators 11 isprovided between the back pressure regulating valve 10 and each of thevapor liquid separators 11 in such a manner that the destination of themobile phase supplied from the back pressure regulating valve 10 can beselected. A check valve 21 for preventing the back flow of a gas fromthe side of the purifying device 13 to each of the vapor liquidseparators 11 is provided between each of the vapor liquid separators 11and the purifying device 13 in correspondence with each of the vaporliquid separators 11.

Each of the pumps 3 and 5 is a pump capable of delivering a constantamount of a liquid. The back pressure regulating valve 10 is a valvethat opens or closes to let the mobile phase having a pressure equal toor higher than a predetermined pressure (for example, 20 MPa) escape tothe side of the back pressure regulating valve 10 close to each of thevapor liquid separators 11, that is, the secondary side of the backpressure regulating valve 10 in such a manner that the pressure of thesystem on the side of the back pressure regulating valve 10 close to thecolumn 8, that is, the primary side of the back pressure regulatingvalve 10 is kept at the predetermined pressure. In addition, the backpressure regulating valve 15 is a valve that opens or closes to let agas having a pressure equal to or higher than a predetermined pressure(for example, 9.5 MPa) escape to the side of the back pressureregulating valve 15 close to the pipe for discharging, that is, thesecondary side of the back pressure regulating valve 15, in such amanner that the pressure on the side of the back pressure regulatingvalve 15 close to each of the purifying device 13 and the heat exchanger2, that is, the primary side of the back pressure regulating valve 15 iskept at the predetermined pressure. In addition, the detector 9 isconnected to a controller (not shown) for controlling the opening andclosing of the predetermined valve 20, the tank 12, and the tank 14 inaccordance with the result of the detection by the detector 9.

As shown in each of FIGS. 2 and 3, each of the vapor liquid separators11 has: a cylindrical external cylinder 31 having ends both of whichbeing opened; a flange portion 32 for clogging an opening at one end ofthe external cylinder 31; an introduction portion 33 as a pipe which isprovided along the tangent of the inner peripheral surface in the crosssectional shape of the external cylinder 31 and which is opened in theexternal cylinder 31; an internal cylinder 34 having ends both of whichbeing opened and which penetrates through the flange portion 32 toextend closer to the other end side of the external cylinder 31 than theintroduction portion 33; a jacket 35 that covers the outer peripheralwall surface of the external cylinder 31 for forming a circulating pathfor a heat medium; and a mobile phase supply pipe 36 that passes theinside of the jacket 35 to be connected to the introduction portion 33.

The jacket 35 is provided with a discharge orifice 38 for the heatmedium on one end side of the jacket 35, and is provided with a supplyorifice 37 for the heat medium on the other end side of the jacket 35.The mobile phase supply pipe 36 is passed from one end portion of thejacket 35 through the inside of the jacket 35, wound on the outerperiphery of the external cylinder 31 from the one end side of thejacket 35 to the other end side of the jacket 35 in a spiral fashion,and taken to the outside from the other end side of the jacket 35 to beconnected to the introduction portion 33. The mobile phase supply pipe36 is connected to the valve 20. The internal cylinder 34 is connectedto the purifying device 13. The other end of the external cylinder 31 isconnected to the tank 12. The supply orifice 37 and the dischargeorifice 38 are connected to heat medium circulating device (not shown).

The purifying device 13 has the same constitution as that of each of thevapor liquid separators 11 shown in each of FIGS. 2 and 3. In thepurifying device 13, the mobile phase supply pipe 36 is connected toeach of the check valves 21. The internal cylinder 34 is connected tothe back pressure regulating valve 15. The other end of the externalcylinder 31 is connected to the tank 14. The supply orifice 37 and thedischarge orifice 38 are connected to heat medium circulating device(not shown) as in the case of each of the vapor liquid separators 11.

A heat medium circulates through the jackets 35 of the vapor liquidseparators 11 and the purifying device 13, whereby the temperature ofeach external cylinder 31 is regulated to a predetermined temperature.

In addition to those components described above, valves such as a valve,a check valve, and a safety valve, various detecting device such as apressure gauge, a thermometer, and a flow meter, and peripheral devicessuch as a heater, a brine chiller, and an accumulator are provided forappropriate sites of the supercritical fluid chromatographicfractionation apparatus although they are not shown.

In this embodiment, the pressure regulating valve 16 is regulated sothat carbon dioxide is supplied from the bomb 1 to the heat exchanger 2at a predetermined pressure (for example, 4 MPa). Carbon dioxide iscooled and liquefied in the heat exchanger 2.

The liquefied gas of carbon dioxide formed in the heat exchanger 2 isstored in the buffer tank 18, and is supplied to the heat exchanger 6 bythe pump 3. The liquefied gas supplied to the heat exchanger 6 issupplied with an organic solvent such as a lower alcohol delivered fromthe solvent tank 4 by the pump 5, whereby the liquefied gas and theorganic solvent are mixed. The mixed solvent is supplied to the heatexchanger 6. In the heat exchanger 6, the mixed solvent is heated sothat the liquefied gas in the mixed solvent is turned into asupercritical fluid. In addition, the temperature of a mobile phaseobtained by mixing the supercritical fluid and the solvent is regulatedto the temperature of the column 8 set by the column oven 19 (forexample, 40° C.). A solution of a separation object is injected as asample from the injector 7 into the mobile phase of which thetemperature has been regulated.

The sample injected from the injector 7 is sent to the column 8, andvarious substances in the sample are divided in association with theirpassage through the column 8.

The substances in the mobile phase that has passed the column 8 aredetected by the detector 9. The predetermined valve 20 is opened and theother valves 20 are closed in accordance with the result of thedetection by the detector 9. The mobile phase that has passed thedetector 9 is sent to the back pressure regulating valve 10. The passingof the mobile phase through the back pressure regulating valve 10reduces the pressure of the mobile phase, and the mobile phase issupplied through the predetermined valve 20 to the mobile phase supplypipe 36 of the predetermined vapor liquid separator 11.

In the vapor liquid separator 11, the mobile phase supplied to themobile phase supply pipe 36 moves in the jacket 35 along the outerperipheral surface of the external cylinder 31 in a spiral fashionbefore it is supplied to the introduction portion 33. Thus, thetemperature of the mobile phase is preliminarily regulated to thetemperature of the external cylinder 31 before the mobile phase issupplied to the introduction portion 33. The mobile phase of which thetemperature has been regulated is introduced from the introductionportion 33 to the external cylinder 31. The mobile phase falls whilecausing a circular motion along the circumferential direction of theinner peripheral wall surface of the external cylinder 31.

In this process, most of carbon dioxide in the mobile phase is dividedas a gas from the mobile phase. The gas divided from the mobile phasepasses the internal cylinder 34 to be discharged from the vapor liquidseparator 11. The solvent remaining as a result of the division of thegas contains a predetermined substance separated from the sample in thecolumn 8. The solvent falls along the inner peripheral wall surface ofthe external cylinder 31 to be stored in the tank 12. The substance inthe solvent stored in the tank 12 is brought into a concentrated statewhen the pressure of the system containing the solvent is releasedbecause liquefied carbon dioxide is released from the solvent so thatthe solubility of the substance in the solvent reduces. After thepressure release, a known method such as vacuum concentration is adoptedas required so that the substance is precipitated out of the solvent, orthe solvent is volatilized. Thus, the substance is taken from thesolvent.

In the external cylinder 31, a droplet of the mobile phase generatesupon introduction of the mobile phase into the external cylinder 31 withstrong momentum or during the flow of the mobile phase in the externalcylinder 31 as described above. The internal cylinder 34 extends closerto the other end side (lower side in the vapor liquid separator 11 ofFIG. 2) of the external cylinder 31 than the introduction portion 33.Accordingly, the droplet is absorbed by the solvent again, or falls asit is along the external cylinder 31, so it hardly enters the internalcylinder 34. Therefore, a gas mixed with a small amount of the dropletis recovered from the internal cylinder 34.

In the purifying device 13, the gas recovered from the internal cylinder34 passes from the mobile phase supply pipe 34 to the introductionportion 33 to be introduced into the external cylinder 31. Then, the gaspasses the internal cylinder to be discharged from the purifying device13. The purifying device 13 removes a trace amount of a solvent mixedinto the gas owing to the droplet from the gas.

The back pressure regulating valve 15 keeps the pressure on the primaryside of the back pressure regulating valve 15 at a predeterminedpressure higher than the pressure set by the pressure regulating valve16. Therefore, the gas discharged from the purifying device 13 issupplied to the heat exchanger 2 so long as the pressure of the gas isequal to or higher than the predetermined pressure. At this time, higherpriority is placed on the supply of the gas purified by the purifyingdevice 13 to the heat exchanger 2 than on the supply of the gas from thebomb 1 to the heat exchanger 2. Therefore, the gas purified by thepurifying device 13 is preferentially used as a liquefied gas. It shouldbe noted that the flow of the gas purified by the purifying device 13 tothe bomb 1 is prevented by the check valve 17.

When the pressure of the gas discharged from the purifying device 13 islower than the pressure set by the pressure regulating valve 16, carbondioxide is supplied from the bomb 1 to the heat exchanger 2. The flow ofthe gas from the bomb 1 to the purifying device 13 is prevented by theback pressure regulating valve 15.

Meanwhile, the back pressure regulating valve 15 supplies an excessivegas for keeping the predetermined pressure to the pipe for discharging.The gas is discharged to outside air directly or after having beensubjected to an appropriate treatment. Alternatively, the gas may becompressed by a compressor or the like to be stored in a bomb andrecycled.

After that, each of the valves 20 is appropriately opened and closed inaccordance with the substance detected by the detector 9, and a desiredsubstance in the sample is taken out.

In this embodiment, the recovery rate of carbon dioxide supplied for usein a supercritical fluid is theoretically about 80% when methanol isused as an organic solvent. Unrecovered carbon dioxide (that is, 20%) isconsidered to be mainly a portion redissolved into the organic solventin each of the vapor liquid separators 11.

In addition, in this embodiment, the use of a supercritical fluid ofcarbon dioxide enables the separation of an acidic substance such as anorganic carboxylic acid as it is with neither addition of an acid to themobile phase nor esterification of the acidic substance. This isprobably because carbon dioxide in the mobile phase shows weak acid. Thedecomposition of the acidic substance as the desired substance can besuppressed because there is no need for adding an acid to the mobilephase. In addition, the recovery rate of the acidic substance from thesample can be increased to be higher than that in the case where theacidic substance is esterified before it is separated because there isno need for esterification.

In addition, in this embodiment, the use of a column having an opticalresolution ability enables separation of an optical isomer at highpurity. For example, in the case of the optical resolution of theracemic body of guaifenesin using a CHIRALCEL OD (1 cmΦ×25 cmL)manufactured by DAICEL CHEMICAL INDUSTRIES, LTD. as the column, an Rbody having an optical purity of 99.0% can be obtained in a yield of94.9%, and an S body having an optical purity of 98.8% can be obtainedin a yield of 98.8%.

In addition, in this embodiment, the intermittent injection of thesample into the mobile phase enables the separation of the desiredsubstance in the sample with high productivity. For example, in the caseof the above-mentioned optical resolution, 4.06 kg of the optical isomercan be produced per day by using 1 kg of a filler for the column.

In this embodiment, after the liquefied gas has been mixed with thesolvent, the temperature of the mixed solvent is regulated, and theliquefied gas in the mixed solvent is turned into the supercriticalfluid. Alternatively, for example, the following procedure may beadopted: a heat exchanger, which heats the liquefied gas before beingmixed with the solvent in such a manner that the temperature of theliquefied gas exceeds a critical temperature, is further provided toform the supercritical fluid, the supercritical fluid is mixed with thesolvent to form a mobile phase, and the temperature of the mobile phaseis regulated to a temperature suitable for separation by the column 8.

In addition, in this embodiment, each of the vapor liquid separators 11is installed to divide the gas upward, and the solvent downward.However, the present invention is not limited to such installation.Alternatively, the vapor liquid separators 11 and the purifying device13 may be installed obliquely or orthogonally to a vertical direction.

In addition, in this embodiment, each of the vapor liquid separators 11is provided with the single tank 12. However, the number of the tanks 12is not particularly limited in the present invention. For example, thefollowing procedure can be adopted: each of the vapor liquid separators11 is provided with multiple tanks connected in series, decompression isperformed between the tanks at any time, and the redissolution of a gasinto the solvent is suppressed. For example, recovering carbon dioxidedissolved into the solvent collected by the tank 12 below each of thevapor liquid separators 11 requires recovering carbon dioxide by usingany one of second and subsequent tanks to be further connected to thetank 12. In such a case, the pressure of each of the second andsubsequent tanks is further reduced (for example, 1 MPa). Carbon dioxidereleased from the solvent in any one of the second and subsequent tankscan be compressed and liquefied by using a compressor, and can be sentto the buffer tank 18.

In this embodiment, the mobile phase containing the desired substance inthe sample is subjected to vapor liquid separation. Accordingly, thesolvent as the divided liquid contains the desired substance, and thedesired substance in a concentrated state can be easily recovered onlyby releasing the divided solvent from a pressure. In addition, thedesired substance in the concentrated state can be recovered only byreleasing the solvent from a pressure, so this embodiment isadvantageous also for the separation of a substance unstable in heatthat is apt to change with time owing to heat.

In addition, in this embodiment, the gas obtained by subjecting themobile phase to vapor liquid separation is supplied to the heatexchanger 2 against the pressure of the gas from the bomb 1, whereby theliquefied gas is formed. Accordingly, carbon dioxide from which thesupercritical fluid is formed can be recycled in supercritical fluidchromatography using a mixed solvent of the supercritical fluid ofcarbon dioxide and an organic solvent as a mobile phase. As a result, acost for carbon dioxide can be reduced. In addition, the discharge ofcarbon dioxide to an environment in supercritical fluid chromatographycan be suppressed.

In addition, in this embodiment, the vapor liquid separators 11 eachprovided with the internal cylinder 34 extending in the externalcylinder 31 up to the position at which no droplet of the mobile phasegenerates in the external cylinder 31 are used. Accordingly, mixing witha droplet of the mobile phase that generates upon vapor liquidseparation can be reduced, and a gas having a high purity that can berecycled as the mobile phase for new chromatography can be obtained bymeans of vapor liquid separation.

In addition, in this embodiment, the mobile phase before beingintroduced into the external cylinder 31 is passed through the inside ofthe jacket 35 for regulating the temperature of the external cylinder31. Accordingly, the temperature of the mobile phase before beingintroduced into the external cylinder 31 can be regulated to thetemperature of the external cylinder 31 in advance. Therefore,efficiency of vapor liquid separation increased by regulating thetemperature of the external cylinder 31 can be further enhanced.

In addition, in this embodiment, the mobile phase supply pipe 36 iswound on the outer peripheral wall surface of the external cylinder 31in the jacket 35. Accordingly, the efficiency of vapor liquid separationincreased by regulating the temperature of the external cylinder 31 canbe further increased by using a compact constitution.

In addition, in this embodiment, the external cylinder 31 is of acylindrical shape, so a pressure is uniformly applied to the peripheralwall of the external cylinder 31. Accordingly, the durability of each ofthe vapor liquid separators 11 in vapor liquid separation under ahigh-pressure condition such as supercritical fluid chromatography canbe further improved.

In addition, in this embodiment, the purifying device 13 is provided, soa trace of an impurity such as the solvent in the gas divided by each ofthe vapor liquid separators 11 is further removed. As a result, a gaswith an improved purity can be recovered, and can be recycled as themobile phase.

In addition, in this embodiment, the check valve 17 for preventing theflow of a gas from the heat exchanger 2 to the bomb 1 is providedbetween the bomb 1 and the heat exchanger 2, so the gas purified by thepurifying device 13 can be prevented from flowing into the bomb 1.

In addition, in this embodiment, the check valve 21 for preventing theflow of a gas from the purifying device 13 to each of the vapor liquidseparators 11 is provided between each of the vapor liquid separators 11and the purifying device 13 in correspondence with each of the vaporliquid separators 11. Accordingly, the back flow of a gas from thepurifying device 13 to the external cylinder 31 or the inflow of a gasfrom one of the vapor liquid separators 11 to any one of the other vaporliquid separators 11 can be prevented.

Industrial Applicability

According to the present invention, in a method of separating asubstance by supercritical fluid chromatography using a mobile phasecomprising a supercritical fluid formed from a liquefied gas obtained byliquefying a gas and a solvent, the mobile phase that has passed acolumn is subjected to vapor liquid separation, and, when the pressureof the gas divided from the mobile phase is higher than the pressure ofa gas supplied for the formation of the liquefied gas from gas supplydevice, the gas divided from the mobile phase is liquefied. Accordingly,a gas from which a supercritical fluid is formed can be recycled in amethod of separating a substance by supercritical fluid chromatographyusing a mobile phase comprising the supercritical fluid and a solvent.

In the present invention, the use of a vapor liquid separator comprisingan external cylinder having ends both of which being opened, a flangeportion for clogging an opening at one end of the external cylinder, anintroduction portion for introducing a mobile phase into the externalcylinder in a circumferential direction along the inner peripheral wallsurface of the external cylinder, and an internal cylinder having endsboth of which being opened and which penetrates through the flangeportion to extend closer to the other end side of the external cylinderthan the introduction portion in the step of dividing the mobile phaseinto a solvent and a gas, is further effective in improving thedurability of the vapor liquid separator and performing vapor liquidseparation with high efficiency in supercritical fluid chromatographyunder a high-pressure condition.

In the present invention, further providing the vapor liquid separatorwith a jacket that covers the outer peripheral wall surface of theexternal cylinder for regulating the temperature of the externalcylinder and a mobile phase supply pipe that passes the inside of thejacket to be connected to the introduction portion for supplying themobile phase to the introduction portion is further effective in furtherimproving the efficiency of vapor liquid separation in supercriticalfluid chromatography.

In the present invention, using a sample which comprises a mixture ofoptical isomers and of which the desired substance is one of the opticalisomers is further effective in efficiently fractionating an opticalisomer with further suppressed mixing with an impurity and having a highpurity.

In addition, the vapor liquid separator of the present invention has theabove-described constitution, so it can be widely used in a separationof a gas with further suppressed mixing with an impurity and having ahigh purity, and in vapor liquid separation in the technical field wheresuch gas is recycled.

In addition, further providing the vapor liquid separator of the presentinvention with the jacket and the mobile phase supply pipe is furthereffective in improving separation efficiency in vapor liquid separation.

The invention claimed is:
 1. A vapor liquid separator for use in amethod of separating a substance by supercritical fluid chromatographyof injecting a sample comprising a desired substance into a mobile phasecomprising a supercritical fluid and a solvent, passing the samplethrough a column for separating the desired substance from the sample toseparate the desired substance in the sample, dividing the mobile phasethat has been passed through the column into the solvent and a gas, andseparating the desired substance from the solvent, wherein the vaporliquid separator comprises: an external cylinder having ends both ofwhich being opened; a flange portion for clogging an opening at one endof the external cylinder; an introduction portion for introducing themobile phase into the external cylinder in a circumferential directionalong an inner peripheral wall surface of the external cylinder; and aninternal cylinder having ends both of which being opened and whichpenetrates through the flange portion to extend closer to an other endside of the external cylinder than the introduction portion.
 2. Thevapor liquid separator according to claim 1, further comprising: ajacket that covers an outer peripheral wall surface of the externalcylinder for regulating a temperature of the external cylinder; and amobile phase supply pipe that passes an inside of the jacket to beconnected to the introduction portion for supplying the mobile phase tothe introduction portion.
 3. The vapor liquid separator according toclaim 1, wherein: the sample comprises a mixture of optical isomers; andthe desired substance comprises one of the optical isomers.
 4. The vaporliquid separator according to claim 2, wherein: the sample comprises amixture of optical isomers; and the desired substance comprises one ofthe optical isomers.